Subcategory: Cancer Research
David Powell - Xavier University of Louisiana
Co-Author(s): Kylar Wiltz, Sruti Chandra, Tarun Mandal, Muniruzzaman Syed, and Anup Kundu, Xavier University, New Orleans, LA
The Multi Drug Resistance gene (MDR) is one of the major factors that allow metastatic breast cancer cells to develop resistance to chemotherapeutic agents such as doxorubicin. Hence, the use of traditional chemotherapeutic agents against these aggressive cancer lines often fail. In addition, the traditional treatment options for metastatic breast cancer have severe side effects. Therefore, the development of an effective therapeutic strategy to circumvent the MDR of metastatic breast cancer is highly anticipated. The MDR of metastatic breast cancer cells was been associated with the overexpression of a Phospho-glycoprotein (P-gp) transporter. Even though the overexpression of P-gp could be minimized by silencing it with siRNA, the question is how can it be selectively targeted to the cancer cells? We propose that if we coat the particles by labeling them with a surface aptamer, these nanoparticles could enhance the selectively and delivery of both siRNA (downregulating the gene responsible for the P-gp transporter) and doxorubicin (killing the targeted cancer cells) into the metastatic breast cancer cells. Our hypothesis is that conjugating nanoparticles with a cancer cell specific aptamer should allow more selective delivery of the therapeutic drugs to tumor cells leading to an enhanced cellular toxicity and antitumor effect, as compared to unconjugated nanoparticles.
The primary objective of this project is to develop a targeted nanocarrier delivery system for siRNA into breast cancer cells. For targeted delivery, Aptamer A6 has been used which can bind to Her-2 receptors on breast cancer cells. In order to bind the aptamer to the particle surface, maleimide-terminated PEGDSPE (MalPEG) was incorporated into the nanoparticles. Initially, 3 different formulations of blank hybrid nanoparticles (i.e. F40, F21, and F31) were prepared by combining different amounts of DOTAP, cholesterol, PLGA or PLGA-PEG and MalPEG in a specific ratio. Next these particles would be compressed into a smaller diameter by using high pressure homogenization (HPH). Then protamine sulphate-condensed the GAPDH siRNA (TRITC conjugated; red) which it then became encapsulated into those nanoparticles. Finally the particles were incubated with aptamer A6 (FITC conjugated; green) for surface labeling. Aptamer labeled-nanoparticles having PLGA are smaller in size than those having PLGA-PEG. Surface charge was reduced when the particles were labeled with the aptamer. Cell transfection was increased significantly when the particles were labeled with the aptamer. Cell transfection was increased significantly in Her2 (+) SKBR-3 cells and Her-2 (+) 4T1-Her-2 cells but not in Her-2 (-) MCF-7 and Her-2 (-) MDA MB-231 cells. No cellular toxicity was observed for any of these formulations. This preliminary study concludes that smaller hybrid particles with PLGA could be further optimized for aptamer mediated targeted delivery of siRNA to breast cancer cells.
References: Bao, L., Haque, A., Jackson, K., Hazari, S., Moroz, K., Jetly, R., and Dash, S. Increased expression of P-glycoprotein is associated with doxorubicin chemoresistance in the metastatic 4T1 breast cancer model. Am J Pathol. 2011, 178(2): 838-52. Bao, L., Hazari, S., Mehra, S., Kaushal, D., Moroz, K., Dash, S. Increased expression of P-glycoprotein and doxorubicin chemoresistance of metastatic breast cancer is regulated by miR -298. Am J Pathol. 2012, 180(6): 2490-503.
Funder Acknowledgement(s): This work was funded in part by LCRC Seed Grant, CUR from Xavier University of Louisiana, BoR SURE Grant, NSF and RCMI Grant (2G12MD007595-06).
Faculty Advisor: Anup Kundu,